title: In vitro phenotyping of recombinant hepatitis B virus containing the polymerase/reverse transcriptase gene from clinical isolates.
abstract: This chapter describes the procedures for the generation of recombinant hepatitis B virus (HBV) that contains the polymerase/reverse transcriptase (pol/RT) gene from clinical isolates. It also contains procedures for phenotypic testing of the recombinant HBV in HepG2 cells for susceptibility to nucleos(t)ide analogs. Amplification and cloning of the pol/RT gene as opposed to the full-length HBV genome from a clinical isolate allows for clinical samples with lower viral loads to be successfully analyzed. Furthermore, it enables a direct comparison of sequence changes in the pol/RT region between isolates.
authors.affiliation: Gilead Sciences Inc., Foster City, CA, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Liu
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: Kitrinos
mesh_terms: D018929:Cell Culture Techniques; D003001:Cloning, Molecular; D004261:DNA Replication; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D015751:Genes, pol; D005822:Genetic Vectors / Q000235:genetics / Q000302:isolation & purification; D056945:Hep G2 Cells; D006515:Hepatitis B virus / Q000302:isolation & purification / Q000502:physiology*; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction; D010641:Phenotype; D010957:Plasmids / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D012194:RNA-Directed DNA Polymerase / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D011995:Recombination, Genetic; D014779:Virus Replication
chemical_list: D004279:DNA, Viral; D012194:RNA-Directed DNA Polymerase